FIGURE 6.

Bone formation in Ankrd11^ncko mice is dysregulated in late embryonic development. (A) Palatal region in E14.5 Ankrd11^ctrl (upper row) and Ankrd11^ncko (lower row) embryos H&E stained for orientation (left column) and immunostained for runt-related transcription factor 2 (Runx2) and Osterix (Sp7) (middle and right columns). Scale bars indicate 100 μm. (B) Quantification of % occupancy of field of view stained for Runx2 and Sp7 (independent two-sample T-test assuming unequal variances). (C) Low and high magnification images of palatal bone in P0 Ankrd11^ctrl (upper row) and Ankrd11^ncko (lower row) pups immunostained for Runx2 (left two columns) and Sp7 (right two columns). DAPI (blue) was used to counterstain nuclei. (D) Representation of individual cells expressing Runx2 and Sp7 generated using ImageJ showing altered cell distribution in the mutant. Red dotted line and arrowhead indicates extended trabeculae in control, whereas red circles and yellow arrowheads indicate more contained cellular clusters in the mutant. Scale bars in first and third columns represent 100 μm, and second and fourth represent 50 μm. (E) Quantification of Runx2 and Sp7 expression at P0 (left: number of positive cells; right: percent area per field of view) showing significant reduction of the area of Runx2 expression, whereas Sp7 expression domain was more contained but not statistically different at P0 (n = minimum of six sections form 3–5 pups/genotype, independent two-sample T-test assuming unequal variances, **p < 0.001).